Study Of The Effect On The Proliferation Of Rat Hepatocytes Cultured With Echinococcus Multilocularis Vesicle Fluid In Vitro

Objective 1.Found the method of the primary culture of rat hepatocytes without serum successfully.2.To explore the effect of Echinococcus multilocularis vesicle fluid (EmF) on liver cell proliferation in vitro.Methods 1.Hepatocytes were isolated from Wistar rat by collagenase digestion and cultured in 35mm culture capsule coated with collagen I at 1×106 . after cultured in a conditioned medium with with serum or not for 24 hours and 48 hours respectively. Compared them through observing the morphologic change and the cell cycle assessed by flow cytometry. 2.After founding the method of the primary culture of rat hepatocytes without serum, hepatocytes were incubated with EMF for 24 hours and 48 hours respectively. The morphologic change of cultured hepatocytes was observed by fluorescence microscope, the cell proliferation of the hepatocytes was assessed by flow cytometry. The toxic effect of EmF was assessed by MTT mothed through co-cultured with 1×105 hepatocytes cultured in 96-well cultured plate which coated with collagen I. Results 1. Found the method of the primary culture of rat hepatocytes without serum: Hepatocytes obtained by collagenase digestion were intact and have a good viability (>90%), which grew well no matter cultured with serum or not. Compare with cultured with serum,the PI of hepatocytes cultured without serum decreased slightly after 24 hours and 48 hours( P>0.05) . 2. Hepatocytes incubated with cyst fluid were intact and have a good function. The MTT assay showed cyst fluid had no cell toxicity on rat hepatocytes. the proliferation index of hepatocytes cultured with cyst fluid for 24 hours decreased remarkably from 82.0%±6.0% to 49.35%±4.0%(P<0.05).the proliferation index of hepatocytes cultured with cyst fluid for 48 hours decreased remarkably from 67%±10% to 27.61%±6%(P<0.05). Conclusion 1.found the method of the primary culture of rat hepatocytes without serum successfully. 2.The EmF had no inhibition on rat hepatocytes cultured in vitro and could inhibit the proliferation of hepatocytes.